Advanced Phytochemical-Based Nanocarrier Systems for the Treatment of Breast Cancer.

As the world's most prevalent cancer, breast cancer imposes a significant societal health burden and is among the leading causes of cancer death in women worldwide. Despite the notable improvements in survival in countries with early detection programs, combined with different modes of treatment to eradicate invasive disease, the current chemotherapy regimen faces significant challenges associated with chemotherapy-induced side effects and the development of drug resistance. Therefore, serious concerns regarding current chemotherapeutics are pressuring researchers to develop alternative therapeutics with better efficacy and safety. Due to their extremely biocompatible nature and efficient destruction of cancer cells via numerous mechanisms, phytochemicals have emerged as one of the attractive alternative therapies for chemotherapeutics to treat breast cancer. Additionally, phytofabricated nanocarriers, whether used alone or in conjunction with other loaded phytotherapeutics or chemotherapeutics, showed promising results in treating breast cancer. In the current review, we emphasize the anticancer activity of phytochemical-instigated nanocarriers and phytochemical-loaded nanocarriers against breast cancer both in vitro and in vivo. Since diverse mechanisms are implicated in the anticancer activity of phytochemicals, a strong emphasis is placed on the anticancer pathways underlying their action. Furthermore, we discuss the selective targeted delivery of phytofabricated nanocarriers to cancer cells and consider research gaps, recent developments, and the druggability of phytoceuticals. Combining phytochemical and chemotherapeutic agents with nanotechnology might have far-reaching impacts in the future.

On the origin of matrix mechanism in protocells and key problems of molecular biology.

The theory of the emergence of the matrix mechanism in protocells on complexes of minerals (apatite, carbonate-apatite, calcite, and quartz) with the reciprocal proportions and with the participation of the gas phase radicals (NH3, CH4, and CO) is considered. The structure of apatite and carbonate-apatite predetermined the formation of a double helix of DNA with the complementary pairs of purine-pyrimidine bases, as well as RNA strands complementary to DNA, and helical protein chains combined into supramolecular structures with RNA. It is proposed that during the Archean Eon, a gradual replacement of the mineral matrix with organic matter took place. The site of the origin of the matrix mechanism is the defect-free and growing defective zone of apatite and carbonate-apatite. The size and specificity of DNA, complementary-bound RNA and protein molecules in supramolecular protein-RNA complexes increased as defects accumulated in the structure of minerals. An increase in the size of RNA transcripts was accompanied by an increase in the number of protein molecules in supramolecular protein-RNA complexes. At the first, anhydrous, stage, the formation of a transcriptional-translational apparatus in the form of a crystalline organic-mineral complex -DNA, RNA and protein, based on the "spiral into spiral" principle of gas phase elements. The appearance of water determined the launch of the transcriptional-translational apparatus and the transformation of the organo-mineral crystalline complex into a liquid-crystalline state. A detailed description of the preparation and launch of the matrix mechanism is given. The following problems are discussed: the origin of ribosomal proteins and the role of super-specific aminoacyl-tRNA synthetase as a true carrier of genetic information; properties of the genetic code and synthesis of protocells without violating the second law of thermodynamics; the origin of biological asymmetry; the appearance of nanobacteria and dark genetic matter of eukaryotic systems.Communicated by Ramaswamy H. Sarma.

Diagnostic and prognostic relevance of CP2c and YY1 expression in hepatocellular carcinoma.

Recent studies have demonstrated an oncogenic role of the transcription factor (TF) CP2c in hepatocellular carcinoma (HCC) based on a strong correlation between CP2c expression, tumor grade, and aggressiveness. We recently found that CP2c directly interacts with another TF, YY1, which is also overexpressed in multiple cancers, including HCC. To evaluate if these proteins are co-regulated in carcinogenesis, we analyzed the expression of CP2c and YY1 in HCC (n = 136) tissues and examined the correlation between their expression and clinicopathological characteristics of HCC. Receiver operating characteristic analysis exhibited the validity of CP2c and nuclear YY1 expression as a diagnostic factor in HCC tissues. High expression of CP2c was significantly correlated with patient age, and higher histological grade, American Joint Committee on Cancer (AJCC) stage, and small and large vessel invasion in HCC tissues, whereas high expression of nuclear YY1 was significantly associated with higher AJCC stage and small vessel invasion. In univariate and multivariate analyses, high expression of CP2c was significantly correlated with disease free survival (DFS), indicating that CP2c expression is an independent prognostic factor for DFS in HCC patients. Patients with high expression of both CP2c and nuclear YY1 usually had a shorter median survival time and worse DFS prognosis than other patients, suggesting that combined detection of CP2c and nuclear YY1 is a useful prognostic marker in HCC patients.

Comparative Analysis of the Antiviral Activity of Camel, Bovine, and Human Lactoperoxidases Against Herpes Simplex Virus Type 1.

Lactoperoxidase is a milk hemoprotein that acts as a non-immunoglobulin protective protein and shows strong antimicrobial activity. Bovine milk contains about 15 and 7 times higher levels of lactoperoxidase than human colustrum and camel milk, respectively. Human, bovine, and camel lactoperoxidases (hLPO, bLPO, and cLPO, respectively) were purified as homogeneous samples with specific activities of 4.2, 61.3, and 8.7 u/mg, respectively. The optimal working pH was 7.5 (hLPO and bLPO) and 6.5 (cLPO), whereas the optimal working temperature for these proteins was 40 °C. The K m of hLPO, cLPO, and bLPO were 17, 16, and 19 mM, and their corresponding V max values were 2, 1.7, and 2.7 µmol/min ml. However, in the presence of H2O2, the K m values were 11 mM for hLPO and cLPO and 20 mM for bLPO, while the corresponding V max values were 1.17 for hLPO and 1.4 µmol/min ml for cLPO and bLPO. All three proteins were able to inhibit the herpes simplex virus type 1 (HSV-1) in Vero cell line model. The relative antiviral activities were proportional to the protein concentrations. The highest anti-HSV-1 activity was exhibited by bLPO that inhibited the HSV particles at a concentration of 0.5 mg/ml with the relative activity of 100%.

Screening the anti infectivity potentials of native N- and C-lobes derived from the camel lactoferrin against hepatitis C virus.

BACKGROUND: Hepatitis C virus (HCV) infection represents a worldwide health threat that still needs efficient protective vaccine and/or effective drug. The traditional medicine, such as camel milk, is heavily used by the large sector of HCV patients to control the infection due to the high cost of the available standard therapy. Camel milk contains lactoferrin, which plays an important and multifunctional role in innate immunity and specific host defense against microbial infection. Continuing the analysis of the effectiveness of camel lactoferrin against HCV, the current study aimed to separate and purify the native N- and C-lobes from the proteolytically cleaved camel lactoferrin (cLF) and to compare their in vitro activities against the HCV infection in Huh7.5 cells in order to determine the most active domain. METHODS: Lactoferrin and its digested N- and C-lobes were purified by Mono S 5/50 GL column and Superdex 200 5/150 column. The purified proteins were assessed through three venues: 1. To inhibit intracellular replication, HCV infected cells were treated with the proteins at different concentrations and time intervals; 2. The proteins were directly incubated with the viral particles (neutralization) and then such neutralized viruses were used to infect cells; 3. The cells were protected with proteins before exposure to the virus. The antiviral potentials of the cLf and its lobes were determined using three techniques: 1. RT-nested PCR, 2. Real-time PCR, and 3. Flow cytometry. RESULTS: N- and C-lobes were purified in two consecutive steps; using Mono-S and Superdex 200 columns. The molecular mass of N- and C-lobes was about 40 kDa. cLF and its lobes could prevent HCV entry into Huh 7.5 cells with activity reached 100% through direct interaction with the virus. The inhibition of intracellular viral replication by N-lobe is 2-fold and 3-fold more effective than that of the cLF and C-lobe, respectively. CONCLUSION: Generated native N- and C-lobes from camel lactoferrin demonstrated a range of noticeably different potentials against HCV cellular infectivity. The anti-HCV activities were sorted as N-lobe > cLf > C-lobe.

A creature with a hundred waggly tails: intrinsically disordered proteins in the ribosome.

Intrinsic disorder (i.e., lack of a unique 3-D structure) is a common phenomenon, and many biologically active proteins are disordered as a whole, or contain long disordered regions. These intrinsically disordered proteins/regions constitute a significant part of all proteomes, and their functional repertoire is complementary to functions of ordered proteins. In fact, intrinsic disorder represents an important driving force for many specific functions. An illustrative example of such disorder-centric functional class is RNA-binding proteins. In this study, we present the results of comprehensive bioinformatics analyses of the abundance and roles of intrinsic disorder in 3,411 ribosomal proteins from 32 species. We show that many ribosomal proteins are intrinsically disordered or hybrid proteins that contain ordered and disordered domains. Predicted globular domains of many ribosomal proteins contain noticeable regions of intrinsic disorder. We also show that disorder in ribosomal proteins has different characteristics compared to other proteins that interact with RNA and DNA including overall abundance, evolutionary conservation, and involvement in protein-protein interactions. Furthermore, intrinsic disorder is not only abundant in the ribosomal proteins, but we demonstrate that it is absolutely necessary for their various functions.

MoRFpred, a computational tool for sequence-based prediction and characterization of short disorder-to-order transitioning binding regions in proteins.

MOTIVATION: Molecular recognition features (MoRFs) are short binding regions located within longer intrinsically disordered regions that bind to protein partners via disorder-to-order transitions. MoRFs are implicated in important processes including signaling and regulation. However, only a limited number of experimentally validated MoRFs is known, which motivates development of computational methods that predict MoRFs from protein chains. RESULTS: We introduce a new MoRF predictor, MoRFpred, which identifies all MoRF types (α, ß, coil and complex). We develop a comprehensive dataset of annotated MoRFs to build and empirically compare our method. MoRFpred utilizes a novel design in which annotations generated by sequence alignment are fused with predictions generated by a Support Vector Machine (SVM), which uses a custom designed set of sequence-derived features. The features provide information about evolutionary profiles, selected physiochemical properties of amino acids, and predicted disorder, solvent accessibility and B-factors. Empirical evaluation on several datasets shows that MoRFpred outperforms related methods: α-MoRF-Pred that predicts α-MoRFs and ANCHOR which finds disordered regions that become ordered when bound to a globular partner. We show that our predicted (new) MoRF regions have non-random sequence similarity with native MoRFs. We use this observation along with the fact that predictions with higher probability are more accurate to identify putative MoRF regions. We also identify a few sequence-derived hallmarks of MoRFs. They are characterized by dips in the disorder predictions and higher hydrophobicity and stability when compared to adjacent (in the chain) residues. AVAILABILITY: http://biomine.ece.ualberta.ca/MoRFpred/; http://biomine.ece.ualberta.ca/MoRFpred/Supplement.pdf.

Analysis of the metabolites in apical area of Allium cepa roots by high resolution NMR spectroscopy method.

Elucidation of the molecular mechanisms determining the formation of various tissues and organs is one of the central problems of cell biology. High-resolution NMR spectroscopy was applied for the analysis of the metabolites produced at the various areas of the apical part of the onion Allium cepa roots. To this end, three samples were extracted from the root apex (the root cap, the meristem region and the cell elongation zone). These samples were noticeably different in the number of mitoses and the sets of metabolites. Furthermore, the complete stasis of the plant roots and tops growth was registered in heavy water. Comparison of the morphological and NMR data revealed their perfect agreement with the cellular processes occurring in the root apex. The root cap sample was characterized by the greatest mitotic activity reflected in the great variability of the chemical compounds extracted from this area, the high level of energy consumption, and the increased synthesis of the phosphocholines needed for the cell fission. Sample containing the cell elongation zone possessed the high sugar content, which is required for the cell-wall growth. Therefore, our data show that high-resolution NMR spectroscopy can be used for the identification of chemical compounds in the various regions of the onion root apical area.

Conformational properties of the SDS-bound state of alpha-synuclein probed by limited proteolysis: unexpected rigidity of the acidic C-terminal tail.

Alpha-synuclein (alpha-syn) is a "natively unfolded" protein constituting the major component of intracellular inclusions in several neurodegenerative disorders. Here, we describe proteolysis experiments conducted on human alpha-syn in the presence of SDS micelles. Our aim was to unravel molecular features of micelle-bound alpha-syn using the limited proteolysis approach. The nonspecific proteases thermolysin and proteinase K, as well as the Glu-specific V8-protease, were used as proteolytic probes. While alpha-syn at neutral pH is easily degraded to a variety of relatively small fragments, in the presence of 10 mM SDS the proteolysis of the protein is rather selective. Complementary fragments 1-111 and 112-140, 1-113 and 114-140, and 1-123 and 124-140 are obtained when thermolysin, proteinase K, and V8 protease, respectively, are used. These results are in line with a conformational model of alpha-syn in which it acquires a folded helical structure in the N-terminal region in its membrane-bound state. At the same time, they indicate that the C-terminal portion of the molecule is rather rigid, as seen in its relative resistance to extensive proteolytic degradation. It is likely that, under the specific experimental conditions of proteolysis in the presence of SDS, the negatively charged C-terminal region can be rigidified by binding a calcium ion, as shown before with intact alpha-syn. In this study, some evidence of calcium binding properties of isolated C-terminal fragments 112-140, 114-140, and 124-140 was obtained by mass spectrometry measurements, since molecular masses for calcium-loaded fragments were obtained. Our results indicate that the C-terminal portion of the membrane-bound alpha-syn is quite rigid and structured, at variance from current models of the membrane-bound protein deduced mostly from NMR. Considering that the aggregation process of alpha-syn is modulated by its C-terminal tail, the results of this study may provide useful insights into the behavior of alpha-syn in a membrane-mimetic environment.